• Vaccine Name: VRP
• Target Pathogen: Clostridium botulinum
• Target Disease: Botulism
• Vaccine Ontology ID: VO_0004078
• Type: Recombinant vector vaccine
• Preparation: Construction begins with the VEE replicon vector, the capsid helper, and the glycoprotein helper, which contains attenuating mutations. Construction, characterization, and assembly of the replicons into VRP for the BoNT/A HC, the anthrax MAT-PA replicon, the MBGV-GP replicon, and the negative control comprising mutagenized staphylococcal enterotoxin B (mSEB) replicon were then prepared. The BoNT/C HC gene was PCR-cloned into the VEE replicon plasmid by using Cla I restriction enzyme recognition sequence—gene specific primers. The VRP titers are expressed as focus-forming units (FFU) where 1 FFU is equivalent to 1 infectious unit (iu) (Lee et al., 2006).
• Virulence: (Lee et al., 2006)
• Description: The development of multi-agent vaccines offers the advantage of eliciting protection against multiple diseases with minimal inoculations over a shorter time span. Formulations consisted of individual Venezuelan equine encephalitis (VEE) virus replicon-vectored vaccines against a bacterial disease (anthrax), a viral disease (Marburg fever), and a toxin-mediated disease (botulism). The individual VEE replicon particles (VRP) expressed mature 83-kDa protective antigen (MAT-PA) from Bacillus anthracis, the glycoprotein (GP) from Marburg virus (MBGV), or the H(C) fragment from botulinum neurotoxin (BoNT H(C)) (Lee et al., 2006).

Mouse Response

• Host Strain: Swiss, CBA/J
• Vaccination Protocol: Swiss and CBA/J mice were inoculated behind the neck with either a single VRP or with a mixture of VRPs at a dose of 10^7 iu of each VRP in 200 μl of PBS on days 0, 35, and 70. The mice were challenged i.p. on d 105 or 164 with 1000 50% median lethal doses (MLD50) of BoNT/A or BoNT/C diluted in PBS containing 0.2% gelatin, respectively. CBA/J mice were challenged s.c. on d 105 with 10 LD50 units (2 × 10^8 spores) of heat-shocked spores of the Sterne strain of B. anthracis. Positive control mice were inoculated with 0.1 ml of anthrax vaccine adsorbed (AVA, Bioport Corp., Lansing, MI) or with 0.1 ml of pentavalent botulinum toxoid vaccine, and negative control mice were inoculated with mSEB VRP, and all were used as controls for the challenges. As a comparison, the dose of AVA or toxoid vaccine administered to humans is 0.5 ml. Sera for ELISA were obtained 2 d before each inoculation and 2 d before challenge.
• Persistence: The quality (i.e. neutralizing verses non-neutralizing antibody activity) of the anti-C/HC antibody response was considerably better than the anti-A/HC antibody response measured for CBA/J mice or that the BoNT/C circulated in the animals for longer periods before entering neurons thus allowing more time for antibody-mediated neutralization of the toxin in the animal after challenge (Lee et al., 2006).
• Side Effects: No adverse reactions or side effects were noted in mice receiving individual or combination VRP vaccines (Lee et al., 2006).
• Efficacy: Inoculation of Swiss mice on d 0, 35, and 70 with 10^7 iu of BoNT/A HC VRP, either alone or in combinations with MAT-PA or MBGV-GP, or with all three VRP vaccines, completely protected the mice from challenge with BoNT/A. CBA/J mice inoculated with BoNT/A HC VRP were 90% protected from a BoNT/A challenge, whereas a mix of BoNT/A HC and MAT-PA VRP poorly protected the animals (2/10 survived) from the same BoNT/A challenge (Lee et al., 2006).
• Description: Increased concerns about the use of biological agents in acts of terrorism and warfare have also increased the need for the rapid development of vaccines against a wide range of bacteria, toxins, and viruses. The NIAID has classified biological organisms and toxins that could be used in bioterrorism and biowarfare as category A, B, or C priority pathogens. In response, current studies involve a multiagent vaccine that utilizes the VEE virus replicon as a vector for BoNT, anthrax, and MBGV, all of which are classified as category A pathogens (Lee et al., 2006).